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1.
Nature ; 596(7873): 531-535, 2021 08.
Artigo em Inglês | MEDLINE | ID: mdl-34433948

RESUMO

Water is one of the most important, yet least understood, liquids in nature. Many anomalous properties of liquid water originate from its well-connected hydrogen bond network1, including unusually efficient vibrational energy redistribution and relaxation2. An accurate description of the ultrafast vibrational motion of water molecules is essential for understanding the nature of hydrogen bonds and many solution-phase chemical reactions. Most existing knowledge of vibrational relaxation in water is built upon ultrafast spectroscopy experiments2-7. However, these experiments cannot directly resolve the motion of the atomic positions and require difficult translation of spectral dynamics into hydrogen bond dynamics. Here, we measure the ultrafast structural response to the excitation of the OH stretching vibration in liquid water with femtosecond temporal and atomic spatial resolution using liquid ultrafast electron scattering. We observed a transient hydrogen bond contraction of roughly 0.04 Å on a timescale of 80 femtoseconds, followed by a thermalization on a timescale of approximately 1 picosecond. Molecular dynamics simulations reveal the need to treat the distribution of the shared proton in the hydrogen bond quantum mechanically to capture the structural dynamics on femtosecond timescales. Our experiment and simulations unveil the intermolecular character of the water vibration preceding the relaxation of the OH stretch.

2.
Nature ; 589(7841): 310-314, 2021 01.
Artigo em Inglês | MEDLINE | ID: mdl-33268896

RESUMO

Photosynthetic reaction centres harvest the energy content of sunlight by transporting electrons across an energy-transducing biological membrane. Here we use time-resolved serial femtosecond crystallography1 using an X-ray free-electron laser2 to observe light-induced structural changes in the photosynthetic reaction centre of Blastochloris viridis on a timescale of picoseconds. Structural perturbations first occur at the special pair of chlorophyll molecules of the photosynthetic reaction centre that are photo-oxidized by light. Electron transfer to the menaquinone acceptor on the opposite side of the membrane induces a movement of this cofactor together with lower amplitude protein rearrangements. These observations reveal how proteins use conformational dynamics to stabilize the charge-separation steps of electron-transfer reactions.


Assuntos
Complexo de Proteínas do Centro de Reação Fotossintética/química , Complexo de Proteínas do Centro de Reação Fotossintética/metabolismo , Bacterioclorofilas/metabolismo , Sítios de Ligação/efeitos dos fármacos , Clorofila/metabolismo , Clorofila/efeitos da radiação , Cristalografia , Citoplasma/metabolismo , Transporte de Elétrons/efeitos dos fármacos , Elétrons , Hyphomicrobiaceae/enzimologia , Hyphomicrobiaceae/metabolismo , Lasers , Modelos Moleculares , Oxirredução/efeitos da radiação , Feofitinas/metabolismo , Complexo de Proteínas do Centro de Reação Fotossintética/efeitos da radiação , Prótons , Ubiquinona/análogos & derivados , Ubiquinona/metabolismo , Vitamina K 2/metabolismo
3.
Nature ; 584(7821): 479-483, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32788728

RESUMO

Lipopolysaccharide (LPS) resides in the outer membrane of Gram-negative bacteria where it is responsible for barrier function1,2. LPS can cause death as a result of septic shock, and its lipid A core is the target of polymyxin antibiotics3,4. Despite the clinical importance of polymyxins and the emergence of multidrug resistant strains5, our understanding of the bacterial factors that regulate LPS biogenesis is incomplete. Here we characterize the inner membrane protein PbgA and report that its depletion attenuates the virulence of Escherichia coli by reducing levels of LPS and outer membrane integrity. In contrast to previous claims that PbgA functions as a cardiolipin transporter6-9, our structural analyses and physiological studies identify a lipid A-binding motif along the periplasmic leaflet of the inner membrane. Synthetic PbgA-derived peptides selectively bind to LPS in vitro and inhibit the growth of diverse Gram-negative bacteria, including polymyxin-resistant strains. Proteomic, genetic and pharmacological experiments uncover a model in which direct periplasmic sensing of LPS by PbgA coordinates the biosynthesis of lipid A by regulating the stability of LpxC, a key cytoplasmic biosynthetic enzyme10-12. In summary, we find that PbgA has an unexpected but essential role in the regulation of LPS biogenesis, presents a new structural basis for the selective recognition of lipids, and provides opportunities for future antibiotic discovery.


Assuntos
Membrana Celular/química , Proteínas de Escherichia coli/química , Proteínas de Escherichia coli/metabolismo , Escherichia coli/química , Escherichia coli/patogenicidade , Lipopolissacarídeos/química , Lipopolissacarídeos/metabolismo , Amidoidrolases/química , Amidoidrolases/metabolismo , Motivos de Aminoácidos , Membrana Externa Bacteriana/química , Membrana Externa Bacteriana/metabolismo , Sítios de Ligação , Membrana Celular/metabolismo , Estabilidade Enzimática , Escherichia coli/citologia , Escherichia coli/efeitos dos fármacos , Genes Essenciais , Hidrolases/química , Hidrolases/metabolismo , Lipídeo A/química , Lipídeo A/metabolismo , Lipopolissacarídeos/biossíntese , Testes de Sensibilidade Microbiana , Viabilidade Microbiana/efeitos dos fármacos , Modelos Moleculares , Fragmentos de Peptídeos/química , Fragmentos de Peptídeos/metabolismo , Fragmentos de Peptídeos/farmacologia , Periplasma/química , Periplasma/metabolismo , Ligação Proteica , Virulência
5.
Langmuir ; 38(42): 12822-12832, 2022 Oct 25.
Artigo em Inglês | MEDLINE | ID: mdl-36220141

RESUMO

Chemical reactions and biological processes are frequently governed by the structure and dynamics of the interface between two liquid phases, but these interfaces are often difficult to study due to the relative abundance of the bulk liquids. Here, we demonstrate a method for generating multilayer thin film stacks of liquids, which we call liquid heterostructures. These free-flowing layered liquid sheets are produced with a microfluidic nozzle that impinges two converging jets of one liquid onto opposite sides of a third jet of another liquid. The resulting sheet consists of two layers of the first liquid enveloping an inner layer of the second liquid. Infrared microscopy, white light reflectivity, and imaging ellipsometry measurements demonstrate that the buried liquid layer has a tunable thickness and displays well-defined liquid-liquid interfaces and that this inner layer can be only tens of nanometers thick. The demonstrated multilayer liquid sheets minimize the amount of bulk liquid relative to their buried interfaces, which makes them ideal targets for spectroscopy and scattering experiments.

6.
Nature ; 539(7627): 43-47, 2016 11 03.
Artigo em Inglês | MEDLINE | ID: mdl-27680699

RESUMO

BinAB is a naturally occurring paracrystalline larvicide distributed worldwide to combat the devastating diseases borne by mosquitoes. These crystals are composed of homologous molecules, BinA and BinB, which play distinct roles in the multi-step intoxication process, transforming from harmless, robust crystals, to soluble protoxin heterodimers, to internalized mature toxin, and finally to toxic oligomeric pores. The small size of the crystals-50 unit cells per edge, on average-has impeded structural characterization by conventional means. Here we report the structure of Lysinibacillus sphaericus BinAB solved de novo by serial-femtosecond crystallography at an X-ray free-electron laser. The structure reveals tyrosine- and carboxylate-mediated contacts acting as pH switches to release soluble protoxin in the alkaline larval midgut. An enormous heterodimeric interface appears to be responsible for anchoring BinA to receptor-bound BinB for co-internalization. Remarkably, this interface is largely composed of propeptides, suggesting that proteolytic maturation would trigger dissociation of the heterodimer and progression to pore formation.


Assuntos
Bacillus/química , Toxinas Bacterianas/química , Culicidae , Inseticidas/química , Larva , Lasers , Animais , Sítios de Ligação , Cristalização , Cristalografia por Raios X , Culicidae/metabolismo , Concentração de Íons de Hidrogênio , Larva/química , Larva/metabolismo , Modelos Moleculares , Multimerização Proteica , Proteólise , Tirosina/química
7.
J Synchrotron Radiat ; 28(Pt 4): 1100-1113, 2021 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-34212873

RESUMO

Determination of electronic structures during chemical reactions remains challenging in studies which involve reactions in the millisecond timescale, toxic chemicals, and/or anaerobic conditions. In this study, a three-dimensionally (3D) microfabricated microfluidic mixer platform that is compatible with time-resolved X-ray absorption and emission spectroscopy (XAS and XES, respectively) is presented. This platform, to initiate reactions and study their progression, mixes a high flow rate (0.50-1.5 ml min-1) sheath stream with a low-flow-rate (5-90 µl min-1) sample stream within a monolithic fused silica chip. The chip geometry enables hydrodynamic focusing of the sample stream in 3D and sample widths as small as 5 µm. The chip is also connected to a polyimide capillary downstream to enable sample stream deceleration, expansion, and X-ray detection. In this capillary, sample widths of 50 µm are demonstrated. Further, convection-diffusion-reaction models of the mixer are presented. The models are experimentally validated using confocal epifluorescence microscopy and XAS/XES measurements of a ferricyanide and ascorbic acid reaction. The models additionally enable prediction of the residence time and residence time uncertainty of reactive species as well as mixing times. Residence times (from initiation of mixing to the point of X-ray detection) during sample stream expansion as small as 2.1 ± 0.3 ms are also demonstrated. Importantly, an exploration of the mixer operational space reveals a theoretical minimum mixing time of 0.91 ms. The proposed platform is applicable to the determination of the electronic structure of conventionally inaccessible reaction intermediates.

8.
Phys Chem Chem Phys ; 23(2): 1308-1316, 2021 Jan 21.
Artigo em Inglês | MEDLINE | ID: mdl-33367391

RESUMO

Electron scattering on liquid samples has been enabled recently by the development of ultrathin liquid sheet technologies. The data treatment of liquid-phase electron scattering has been mostly reliant on methodologies developed for gas electron diffraction, in which theoretical inputs and empirical fittings are often needed to account for the atomic form factor and remove the inelastic scattering background. In this work, we present an alternative data treatment method that is able to retrieve the radial distribution of all the charged particle pairs without the need of either theoretical inputs or empirical fittings. The merits of this new method are illustrated through the retrieval of real-space molecular structure from experimental electron scattering patterns of liquid water, carbon tetrachloride, chloroform, and dichloromethane.

9.
Anal Chem ; 92(1): 1565-1573, 2020 01 07.
Artigo em Inglês | MEDLINE | ID: mdl-31790200

RESUMO

The method of X-ray footprinting and mass spectrometry (XFMS) on large protein assemblies and membrane protein samples requires high flux density to overcome the hydroxyl radical scavenging reactions produced by the buffer constituents and the total protein content. Previously, we successfully developed microsecond XFMS using microfluidic capillary flow and a microfocused broadband X-ray source at the Advanced Light Source synchrotron beamlines, but the excessive radiation damage incurred when using capillaries prevented the full usage of a high-flux density beam. Here we present another significant advance for the XFMS method: the instrumentation of a liquid injection jet to deliver container free samples to the X-ray beam. Our preliminary experiments with a liquid jet at a bending magnet X-ray beamline demonstrate the feasibility of the approach and show a significant improvement in the effective dose for both the Alexa fluorescence assay and protein samples compared to conventional capillary flow methods. The combination of precisely controlled high dose delivery, shorter exposure times, and elimination of radiation damage due to capillary effects significantly increases the signal quality of the hydroxyl radical modification products and the dose-response data. This new approach is the first application of container free sample handling for XFMS and opens up the method for even further advances, such as high-quality microsecond time-resolved XFMS studies.


Assuntos
Radical Hidroxila/análise , Proteínas de Membrana/análise , Pegadas de Proteínas , Síncrotrons , Corantes Fluorescentes/química , Raios X
10.
Proc Natl Acad Sci U S A ; 114(9): 2247-2252, 2017 02 28.
Artigo em Inglês | MEDLINE | ID: mdl-28202732

RESUMO

To understand how molecules function in biological systems, new methods are required to obtain atomic resolution structures from biological material under physiological conditions. Intense femtosecond-duration pulses from X-ray free-electron lasers (XFELs) can outrun most damage processes, vastly increasing the tolerable dose before the specimen is destroyed. This in turn allows structure determination from crystals much smaller and more radiation sensitive than previously considered possible, allowing data collection from room temperature structures and avoiding structural changes due to cooling. Regardless, high-resolution structures obtained from XFEL data mostly use crystals far larger than 1 µm3 in volume, whereas the X-ray beam is often attenuated to protect the detector from damage caused by intense Bragg spots. Here, we describe the 2 Å resolution structure of native nanocrystalline granulovirus occlusion bodies (OBs) that are less than 0.016 µm3 in volume using the full power of the Linac Coherent Light Source (LCLS) and a dose up to 1.3 GGy per crystal. The crystalline shell of granulovirus OBs consists, on average, of about 9,000 unit cells, representing the smallest protein crystals to yield a high-resolution structure by X-ray crystallography to date. The XFEL structure shows little to no evidence of radiation damage and is more complete than a model determined using synchrotron data from recombinantly produced, much larger, cryocooled granulovirus granulin microcrystals. Our measurements suggest that it should be possible, under ideal experimental conditions, to obtain data from protein crystals with only 100 unit cells in volume using currently available XFELs and suggest that single-molecule imaging of individual biomolecules could almost be within reach.


Assuntos
Cristalografia/métodos , Elétrons , Granulovirus/ultraestrutura , Peptídeos e Proteínas de Sinalização Intercelular/química , Lasers , Cristalografia/instrumentação , Granulovirus/química , Modelos Moleculares , Progranulinas , Estrutura Secundária de Proteína , Síncrotrons
11.
Phys Rev Lett ; 120(13): 133203, 2018 Mar 30.
Artigo em Inglês | MEDLINE | ID: mdl-29694162

RESUMO

We report the observation and analysis of the gain curve of amplified Kα x-ray emission from solutions of Mn(II) and Mn(VII) complexes using an x-ray free electron laser to create the 1s core-hole population inversion. We find spectra at amplification levels extending over 4 orders of magnitude until saturation. We observe bandwidths below the Mn 1s core-hole lifetime broadening in the onset of the stimulated emission. In the exponential amplification regime the resolution corrected spectral width of ∼1.7 eV FWHM is constant over 3 orders of magnitude, pointing to the buildup of transform limited pulses of ∼1 fs duration. Driving the amplification into saturation leads to broadening and a shift of the line. Importantly, the chemical sensitivity of the stimulated x-ray emission to the Mn oxidation state is preserved at power densities of ∼10^{20} W/cm^{2} for the incoming x-ray pulses. Differences in signal sensitivity and spectral information compared to conventional (spontaneous) x-ray emission spectroscopy are discussed. Our findings build a baseline for nonlinear x-ray spectroscopy for a wide range of transition metal complexes in inorganic chemistry, catalysis, and materials science.

12.
Nat Methods ; 11(9): 923-6, 2014 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-25108686

RESUMO

We describe a method to measure ultrafast protein structural changes using time-resolved wide-angle X-ray scattering at an X-ray free-electron laser. We demonstrated this approach using multiphoton excitation of the Blastochloris viridis photosynthetic reaction center, observing an ultrafast global conformational change that arises within picoseconds and precedes the propagation of heat through the protein. This provides direct structural evidence for a 'protein quake': the hypothesis that proteins rapidly dissipate energy through quake-like structural motions.


Assuntos
Transferência de Energia/efeitos da radiação , Lasers , Ficobiliproteínas/efeitos da radiação , Ficobiliproteínas/ultraestrutura , Espalhamento a Baixo Ângulo , Difração de Raios X/métodos , Ficobiliproteínas/química , Conformação Proteica/efeitos da radiação , Doses de Radiação
13.
Nature ; 470(7332): 73-7, 2011 Feb 03.
Artigo em Inglês | MEDLINE | ID: mdl-21293373

RESUMO

X-ray crystallography provides the vast majority of macromolecular structures, but the success of the method relies on growing crystals of sufficient size. In conventional measurements, the necessary increase in X-ray dose to record data from crystals that are too small leads to extensive damage before a diffraction signal can be recorded. It is particularly challenging to obtain large, well-diffracting crystals of membrane proteins, for which fewer than 300 unique structures have been determined despite their importance in all living cells. Here we present a method for structure determination where single-crystal X-ray diffraction 'snapshots' are collected from a fully hydrated stream of nanocrystals using femtosecond pulses from a hard-X-ray free-electron laser, the Linac Coherent Light Source. We prove this concept with nanocrystals of photosystem I, one of the largest membrane protein complexes. More than 3,000,000 diffraction patterns were collected in this study, and a three-dimensional data set was assembled from individual photosystem I nanocrystals (∼200 nm to 2 µm in size). We mitigate the problem of radiation damage in crystallography by using pulses briefer than the timescale of most damage processes. This offers a new approach to structure determination of macromolecules that do not yield crystals of sufficient size for studies using conventional radiation sources or are particularly sensitive to radiation damage.


Assuntos
Cristalografia por Raios X/métodos , Nanopartículas/química , Nanotecnologia/métodos , Complexo de Proteína do Fotossistema I/química , Cristalografia por Raios X/instrumentação , Lasers , Modelos Moleculares , Nanotecnologia/instrumentação , Conformação Proteica , Fatores de Tempo , Raios X
14.
Nature ; 470(7332): 78-81, 2011 Feb 03.
Artigo em Inglês | MEDLINE | ID: mdl-21293374

RESUMO

X-ray lasers offer new capabilities in understanding the structure of biological systems, complex materials and matter under extreme conditions. Very short and extremely bright, coherent X-ray pulses can be used to outrun key damage processes and obtain a single diffraction pattern from a large macromolecule, a virus or a cell before the sample explodes and turns into plasma. The continuous diffraction pattern of non-crystalline objects permits oversampling and direct phase retrieval. Here we show that high-quality diffraction data can be obtained with a single X-ray pulse from a non-crystalline biological sample, a single mimivirus particle, which was injected into the pulsed beam of a hard-X-ray free-electron laser, the Linac Coherent Light Source. Calculations indicate that the energy deposited into the virus by the pulse heated the particle to over 100,000 K after the pulse had left the sample. The reconstructed exit wavefront (image) yielded 32-nm full-period resolution in a single exposure and showed no measurable damage. The reconstruction indicates inhomogeneous arrangement of dense material inside the virion. We expect that significantly higher resolutions will be achieved in such experiments with shorter and brighter photon pulses focused to a smaller area. The resolution in such experiments can be further extended for samples available in multiple identical copies.


Assuntos
Mimiviridae/química , Difração de Raios X/instrumentação , Difração de Raios X/métodos , Elétrons , Temperatura Alta , Lasers , Fótons , Fatores de Tempo , Raios X
15.
Proc Natl Acad Sci U S A ; 111(35): 12769-74, 2014 Sep 02.
Artigo em Inglês | MEDLINE | ID: mdl-25136092

RESUMO

It has long been known that toxins produced by Bacillus thuringiensis (Bt) are stored in the bacterial cells in crystalline form. Here we describe the structure determination of the Cry3A toxin found naturally crystallized within Bt cells. When whole Bt cells were streamed into an X-ray free-electron laser beam we found that scattering from other cell components did not obscure diffraction from the crystals. The resolution limits of the best diffraction images collected from cells were the same as from isolated crystals. The integrity of the cells at the moment of diffraction is unclear; however, given the short time (∼ 5 µs) between exiting the injector to intersecting with the X-ray beam, our result is a 2.9-Å-resolution structure of a crystalline protein as it exists in a living cell. The study suggests that authentic in vivo diffraction studies can produce atomic-level structural information.


Assuntos
Bacillus thuringiensis/química , Proteínas de Bactérias/química , Cristalografia por Raios X/métodos , Endotoxinas/química , Proteínas Hemolisinas/química , Esporos Bacterianos/química , Bacillus thuringiensis/ultraestrutura , Toxinas de Bacillus thuringiensis , Cristalização , Cristalografia por Raios X/instrumentação , Lasers , Esporos Bacterianos/ultraestrutura , Síncrotrons , Difração de Raios X
16.
Int J Mol Sci ; 18(9)2017 Sep 07.
Artigo em Inglês | MEDLINE | ID: mdl-28880248

RESUMO

The photochromic fluorescent protein Skylan-NS (Nonlinear Structured illumination variant mEos3.1H62L) is a reversibly photoswitchable fluorescent protein which has an unilluminated/ground state with an anionic and cis chromophore conformation and high fluorescence quantum yield. Photo-conversion with illumination at 515 nm generates a meta-stable intermediate with neutral trans-chromophore structure that has a 4 h lifetime. We present X-ray crystal structures of the cis (on) state at 1.9 Angstrom resolution and the trans (off) state at a limiting resolution of 1.55 Angstrom from serial femtosecond crystallography experiments conducted at SPring-8 Angstrom Compact Free Electron Laser (SACLA) at 7.0 keV and 10.5 keV, and at Linac Coherent Light Source (LCLS) at 9.5 keV. We present a comparison of the data reduction and structure determination statistics for the two facilities which differ in flux, beam characteristics and detector technologies. Furthermore, a comparison of droplet on demand, grease injection and Gas Dynamic Virtual Nozzle (GDVN) injection shows no significant differences in limiting resolution. The photoconversion of the on- to the off-state includes both internal and surface exposed protein structural changes, occurring in regions that lack crystal contacts in the orthorhombic crystal form.


Assuntos
Cristalografia por Raios X/métodos , Lasers , Proteínas Luminescentes/química , Conformação Proteica , Temperatura
17.
Arch Biochem Biophys ; 602: 61-68, 2016 Jul 15.
Artigo em Inglês | MEDLINE | ID: mdl-26944553

RESUMO

Serial femtosecond crystallography (SFX) employing high-intensity X-ray free-electron laser (XFEL) sources has enabled structural studies on microcrystalline protein samples at non-cryogenic temperatures. However, the identification and optimization of conditions that produce well diffracting microcrystals remains an experimental challenge. Here, we report parallel SFX and transmission electron microscopy (TEM) experiments using fragmented microcrystals of wild type (WT) homoprotocatechuate 2,3-dioxygenase (HPCD) and an active site variant (H200Q). Despite identical crystallization conditions and morphology, as well as similar crystal size and density, the indexing efficiency of the diffraction data collected using the H200Q variant sample was over 7-fold higher compared to the diffraction results obtained using the WT sample. TEM analysis revealed an abundance of protein aggregates, crystal conglomerates and a smaller population of highly ordered lattices in the WT sample as compared to the H200Q variant sample. While not reported herein, the 1.75 Å resolution structure of the H200Q variant was determined from ∼16 min of beam time, demonstrating the utility of TEM analysis in evaluating sample monodispersity and lattice quality, parameters critical to the efficiency of SFX experiments.


Assuntos
Cristalização/métodos , Cristalografia/métodos , Teste de Materiais/métodos , Microscopia Eletrônica de Transmissão/métodos , Proteínas/química , Proteínas/ultraestrutura , Cristalografia/tendências
18.
Nat Methods ; 9(3): 263-5, 2012 Jan 29.
Artigo em Inglês | MEDLINE | ID: mdl-22286383

RESUMO

X-ray free electron laser (X-FEL)-based serial femtosecond crystallography is an emerging method with potential to rapidly advance the challenging field of membrane protein structural biology. Here we recorded interpretable diffraction data from micrometer-sized lipidic sponge phase crystals of the Blastochloris viridis photosynthetic reaction center delivered into an X-FEL beam using a sponge phase micro-jet.


Assuntos
Cristalografia por Raios X/métodos , Bicamadas Lipídicas/química , Proteínas de Membrana/química , Proteínas de Membrana/ultraestrutura , Ligação Proteica , Conformação Proteica/efeitos da radiação , Raios X
19.
Nat Methods ; 9(3): 259-62, 2012 Jan 29.
Artigo em Inglês | MEDLINE | ID: mdl-22286384

RESUMO

Protein crystallization in cells has been observed several times in nature. However, owing to their small size these crystals have not yet been used for X-ray crystallographic analysis. We prepared nano-sized in vivo-grown crystals of Trypanosoma brucei enzymes and applied the emerging method of free-electron laser-based serial femtosecond crystallography to record interpretable diffraction data. This combined approach will open new opportunities in structural systems biology.


Assuntos
Cristalografia por Raios X/métodos , Cristalografia/métodos , Proteínas/química , Proteínas/ultraestrutura , Ligação Proteica/efeitos da radiação , Conformação Proteica/efeitos da radiação , Proteínas/efeitos da radiação , Solubilidade/efeitos da radiação , Raios X
20.
Phys Rev Lett ; 114(9): 098102, 2015 Mar 06.
Artigo em Inglês | MEDLINE | ID: mdl-25793853

RESUMO

We present a proof-of-concept three-dimensional reconstruction of the giant mimivirus particle from experimentally measured diffraction patterns from an x-ray free-electron laser. Three-dimensional imaging requires the assembly of many two-dimensional patterns into an internally consistent Fourier volume. Since each particle is randomly oriented when exposed to the x-ray pulse, relative orientations have to be retrieved from the diffraction data alone. We achieve this with a modified version of the expand, maximize and compress algorithm and validate our result using new methods.


Assuntos
Imageamento Tridimensional/métodos , Mimiviridae/ultraestrutura , Difração de Raios X/métodos , Algoritmos , Elétrons , Lasers , Difração de Raios X/instrumentação
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